Efficacy of antibacterial activity of garlic cloves from Tamil Nadu and Jowai region

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1 International Journal of Research in Medical Sciences Anitha M et al. Int J Res Med Sci May;2(2): pissn eissn Research Article DOI: / ijrms Efficacy of antibacterial activity of garlic cloves from Tamil Nadu and Jowai region M. Anitha*, J. Hemapriya Department of Microbiology, Shri Sathya Sai Medical College & Research Institute, Thiruporur, Sri Balaji Vidyapeeth University, Tamil Nadu, India Received: 26 January 2014 Accepted: 2 February 2014 *Correspondence: M. Anitha, animalar03@gmail.com 2014 Anitha M et al. This is an open-access article distributed under the terms of the Creative Commons Attribution Non-Commercial License, which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited. ABSTRACT Background: The development of antibiotic resistance has become a global health challenge which is causing ineffectiveness of the available antibacterial agents leading to increase in diseases and death rate. Therefore this study intends to investigate the antibacterial action of Aqueous Garlic Extract (AGE) against 9 multidrug-resistant grampositive and gram-negative bacterial isolates, including Staphylococcus aureus, Enterococcus species, ß hemolytic streptococcus, Pseudomonas aeruginosa, Proteus mirabilis, Klebsiella pneumoniae, Escherichia coli, Vibrio cholerae and Serretia marscenes. Methods: Antibacterial activity of different concentrations of AGE by well-diffusion method was recorded by measuring the diameter of zone of inhibition. The Tamil Nadu garlic cloves as well as Jowai region garlic cloves showed antibacterial activity against both GPC and GNR. Results: The maximum zone of inhibition was observed in Tamil Nadu garlic than that of Jowai region, but the only bacteria which showed a better zone of inhibition with Jowai region than Tamil Nadu garlic was Pseudomonas aeruginosa. Conclusion: Thus our study reveals that garlic not only makes the food more spicy & edible with its flavour but can also be used as an effective antibacterial agents against MDR gram positive & gram negative bacteria. Keywords: Bacterial isolates, Antibacterial activity, Tamil Nadu garlic and Jowai region garlic extracts INTRODUCTION The microbial infections are the major cause of morbidity and mortality in the developed and developing country, although a number of antimicrobial agents are available for the treatment and management of infectious diseases. In addition, misuse of the antibiotics which can lead to the development of antibiotic resistance is also a major health concern. 1 Therefore, there is a perpetual need to exploit new bioactive principles with high safety index. Historically, medicinal plants have been a source of novel drug compounds. Plants derived products have made large contributions to human health and well-being. Green pharmacy may become the base for the development of medicines by providing a pharmacophore which could be used for the development of new drug with novel mechanisms of action. 2 A number of medicinal plants have been screened for the antimicrobial activity in recent years 3 and efforts has been taken to identify their active constituents. 4 Garlic is one of the edible plants which have generated a lot of interest throughout human history as a medicinal panacea. A wide range of microorganisms including bacteria, fungi, protozoa and viruses has been shown to International Journal of Research in Medical Sciences April-June 2014 Vol 2 Issue 2 Page 434

2 be sensitive to crushed garlic preparations. Moreover, garlic has been reported to reduce blood lipids and has anti-cancer effects. Chemical analyses of garlic cloves have revealed an unusual concentration of sulphurcontaining compounds (1 3%). 5,6 A daily dose of 1 ml/kg body weight of garlic extract for six months can result in significant reduction in oxidant (free radical) stress in the blood of patients with atherosclerosis and cholesterol circulating in the bloodstream. Garlic (Allium sativum) is a common spice used for flavoring and has been traditionally popular with strong folkloric awareness. It is the edible bulb of lily family, Liliaceae. It contains aromatic sulphur based compounds, which contribute to the characteristic odour and taste. Antimicrobial activity of garlic is attributed to its key component allicin, which is a volatile & active ingredient; it acts by partial inhibition of DNA and protein synthesis and also total inhibition of RNA synthesis as a primary target. 7 The main advantage of using the herbal antimicrobial drug is that the side effects are reduced: The side effects mainly include damage to the normal intestinal flora, bone marrow depression, dysentery, local inflammation and damage to liver. 8 The present study was aimed at determining the in vitro antibacterial activity between TN and Jowai region garlic clove extracts on the isolates of gram-positive and gramnegative organisms with the view to find an alternative means of treating infections caused by them. METHODS Microorganism 9 different strains of gram positive & gram negative organisms: Staphylococcus aureus, Enterococcus species, ß hemolytic streptococcus, Pseudomonas aeruginosa, Proteus mirabilis, Klebsiella pneumoniae, Escherichia coli, Vibrio cholerae and Serretia marscenes cultures were collected from our hospital Shri Sathya Sai Medical College And Research Institute, Thiruporur. The cultures were streaked onto individual agar plates and incubated at 37 C for 24 h. Preparation of aqueous garlic extract (AGE) The Tamil Nadu garlic cloves were obtained from the local market & Jowai region garlic cloves were obtained from the North East region. The spices were cleaned and washed using sterile distilled water. 1 gram of both types of garlic was homogenized using a sterile blender and the extract was sieved using sterile cheese cloth. The concentrations of 100 µl, 50 µl and 25 µl were prepared by mixing with appropriate volumes of sterile distilled water. 9 Antibacterial activity The antibacterial properties of both Tamil Nadu and Jowai garlic cloves were studied by agar well diffusion method and disk diffusion methods. 10 Agar well diffusion method The selected bacterial strains were inoculated into 10 ml of sterile nutrient broth and incubated at 37 C for h. Using a sterile cotton swab, the nutrient broth cultures was swabbed on the surface of sterile agar plates. Agar wells were prepared with the help of sterilized cork borer with 6 mm diameter. Using a micropipette, concentrations of 100 μl, 50 μl, and 25 μl solution of garlic were added to different wells in the plates. The plates were incubated in an upright position at 37 C for 24 h. The diameters of inhibition zone (in mm) were measured. Antibiotic sensitivity testing The microorganisms were also tested for their sensitivity against the antibiotics ciprofloxacin, vancomycin, gentamicin, linezolid, amoxiclav by disc diffusion method. The cultures of test microorganism were enriched in sterile nutrient broth for 16 h at 37 C. Sterile cotton swabs were used to transfer the test cultures aseptically on the surface of sterile nutrient agar plates. Flamed forceps was used to place the antibiotic disc aseptically over the seeded agar plates. The plates were incubated at 37 C for 24 h and the diameters of inhibition zones were measured. Antimicrobial susceptibility was tested by the disk diffusion method according to the Clinical and Laboratory Standards Institute (CLSI) guidelines (CLSI, 2007). 11 RESULTS The antibacterial activity of Tamil Nadu garlic was compared with the antibacterial activity of Jowai region garlic with 3 GPC and 6 GNR which are listed in Table 1. Table 1: Comparison of the antibacterial activity of Tamil Nadu garlic with Jowai region garlic with 3 GPC and 6 GNR. GNR Pseudomonas aeruginosa Proteus mirabilis Klebsiella pneumoniae Escherichia coli Vibrio cholerae Serretia marscenes GPC Staphylococcus aureus Enterococcus species ß hemolytic streptococcus The Tamil Nadu garlic as well as Jowai region garlic showed antibacterial activity against both GPC and GNR. All the 3 GPC exhibited high sensitivity zone with 100 µl of Tamil Nadu garlic (Table 3) along with Vibrio cholerae which is GNR (Table 2). International Journal of Research in Medical Sciences April-June 2014 Vol 2 Issue 2 Page 435

3 Table 2: Antibacterial activity of Tamil Nadu and Jowai region garlic - gram negative bacilli. Tamil Nadu garlic zone of inhibition (mm) Jowai region garlic zone of inhibition (mm) Organisms isolated Negative Negative Pseudomonas aeruginosa NZ 20mm 18mm 16mm NZ 25mm 20mm 19mm Proteus mirabilis NZ 28mm 25mm 20mm NZ 20mm 17mm 15mm Klebsiella pneumoniae NZ 30mm 22mm 20mm NZ 16mm 15mm 14mm Escherichia coli NZ 28mm 26mm 25mm NZ 21mm 20mm 19mm Vibrio cholerae NZ 32mm 30mm 29mm NZ 23mm 16mm 15mm Serretia marscenes NZ 26mm 24mm 20mm NZ 18mm 16mm 15mm NZ - No zone Table 3: Antibacterial activity of Tamil Nadu and Jowai region garlic - gram positive cocci. Tamil Nadu garlic zone of inhibition (mm) Jowai region garlic zone of inhibition (mm) Organisms isolated Negative Negative Staphylococcus aureus NZ 32mm 30mm 20mm NZ 25mm 23mm 20mm Enterococcus species NZ 30mm 23mm 20mm NZ 25mm 20mm 15mm ß hemolytic streptococcus NZ 27mm 26mm 25mm NZ 24mm 18mm 16mm NZ - No zone The 50 µl extract of Tamil Nadu garlic zone showed a better zone in both GPC and GNB isolates but maximum diameter was obtained in Staphylococcus aureus and V. cholerae-30mm, followed by ß hemolytic streptococcus and Escherichia coli-26mm. The 25 µl concentration of the garlic gave best results for V. cholera-29mm, followed by ß hemolytic streptococcus and Escherichia coli-25mm. These zones are less than the results obtained in the previous study: In vitro antibacterial activity and sensitivity of garlic extract at different ph and temperature 12 for Staphylococcus aureus, Escherichia coli, Klebsiella pneumoniae and Pseudomonas aeruginosa. But the 50 µl and 25 µl concentration showed a better result in our study, when compared to the previous study which are listed below in Table 4. Table 4: Antibacterial activity of previous study. Diameter of inhibition zone (mm) Test organisms of different concentrations 100% 50% 25% 10% Escherichia coli Klebsiella pneumoniae Pseudomonas aeruginosa Staphylococcus aureus The Jowai region garlic zone showed a strong positive zone of inhibition of 25mm in GPC namely Staphylococcus aureus, Enterococcus species where as GNB like Escherichia coli showed 21mm, followed by 24mm in ß hemolytic streptococcus and V. cholerae, exhibiting 23mm while Pseudomonas aeruginosa had 25mm with 100 µl extract of Jowai region garlic. Jowai region garlic showed 20mm zone of diameter with other Gram negative bacteria like Proteus mirabilis, Serratia marscenes had 18mm and 16mm with Klebsiella pneumoniae in 100 µl (Figure 4). The lowest diameter of 14mm was seen in Klebsiella pneumoniae for 25 µl concentration of Jowai region garlic. The results obtained from our study, clearly showed that all the bacterial strains which are resistant to antibiotics (Ciprofloxacin, vancomycin, gentamicin, linezolid, amoxiclav) were highly susceptible to garlic clove extracts. It is evident that the Tamil Nadu and Jowai region garlic clove extracts showed better and broader spectrum activity when compared to the antibiotics (Figure 2). Figure 1: Showing Tamil Nadu and Jowai region garlic. International Journal of Research in Medical Sciences April-June 2014 Vol 2 Issue 2 Page 436

4 The Jowai region garlic extract had its maximum zone of inhibition with all the gram positive cocci: Staphylococcus aureus, Enterococcus species and ß hemolytic streptococcus (Table 3) (Figure 6). Figure 2: Kirby Bauer s disk diffusion technique with conventional antibiotic disc, showing with resistance to GNB & GPC. Figure 3: Antibacterial activity of Tamil Nadu garlic by well diffusion method for GNB. Figure 4: Antibacterial activity of Jowai region garlic by well diffusion method for GNB. Figure 5: Antibacterial activity of Tamil Nadu garlic by well diffusion method for GPC. Among the gram negative bacilli again maximum diameter was seen with 100 µl of Jowai region garlic extract in Vibrio cholerae which had 23mm, and Pseudomonas aeruginosa 25mm. These two GNR exhibited the maximum diameter zone (Table 2) (Figure 4) followed by Staphylococcus aureus, Enterococcus faecalis which showed 25mm in 100 µl (Table 3). The only bacteria which showed a better zone of inhibition with Jowai region than Tamil Nadu garlic was Pseudomonas aeruginosa (Table 2). CONCLUSION A total number of 9 isolates were processed for susceptibility testing with aqueous extraction of Tamil Nadu and Jowai region garlic cloves, with the concentration of 1 gram by well diffusion technique as per standard protocol. It was found out that maximum zone of inhibition was observed in Staphylococcus aureus and Vibrio cholerae with Tamil Nadu garlic & Jowai region garlic cloves. Our study is the first of its kind to analyze the antibacterial activity of the Jowai region garlic cloves which has been proved to have a strong sensitive pattern against both Gram positive & Gram negative bacteria as shown in our analysis. Therefore the effectiveness of either the Tamil Nadu or Jowai region garlic cloves is proved beyond doubt that it not only has edible quality in preparing mouth-watering spicy food but also antibacterial activity against both GPC & GNB. ACKNOWLEDGEMENTS The authors are thankful to Dr. Karthika Jayakumar, Professor & Head, Microbiology from Shri Sathya Sai medical college and research institute and Dr. Velmurugan, Professor & Head, CAS in crystallography and biophysics from university of Madras, Guindy campus for their encouragement to carry out this work. Figure 6: Antibacterial activity of Jowai region garlic by well diffusion method for GPC. DISCUSSION The maximum zone of inhibition with Tamil Nadu garlic was observed in GNR, V. cholerae in all the three concentrations (100 µl, 50µL, 25 µl) (Figure 3). In the GPC, Staphylococcus aureus exhibited maximum diameter with 32mm, 30mm, 20mm with 100 µl, 50µL, 25 µl concentrations (Figure 5). Funding: No funding sources Conflict of interest: None declared Ethical approval: Not required REFERENCES 1. Al-Bari MAA, Khan A, Islam MR, Kudrat-E-Zahan E, Rahman MMS, Ul-Islam MA et al. Isolation and in vitro antimicrobial activities of ethyl acetate extract from Streptomyces bangladeshiensis. Res J Microbiol. 2007;2: Menghani E, Pareek A, Negi RS, Ojha CK. Search for antimicrobial potential from certain Indian International Journal of Research in Medical Sciences April-June 2014 Vol 2 Issue 2 Page 437

5 medicinal plants. Res J Med Plants. 2011;5: Premanath R, Sudisha J, Lakshmi Devi N, Aradhya SM. Antibacterial and antioxidant activities of Fenugreek (Trigonella foenum graecum L.) leaves. Res J Med Plants. 2011;5: Tijjani MB, Bello IA, Aliyu AB, Olurishe T, Maidawa SM, Habila JD et al. Phytochemical and antibacterial studies of root extract of Cochlospermum tinctorium A. rich. (Cochlospermacease) Res J Med Plants. 2009;3: Darbyshire B., Henry R. J., Differences in fructan content and synthesis in some Allium species. New Phytol. 1981;87: Koch H. P., Lawson L. D. Garlic, the science and therapeutic application of Allium sativum L. and related species. In: Retford D. C., eds. Lippincott Williams & Wilkins. Baltimore: Wolters Kluwer; 1996: Alli JA, Boboye BE, Okonko IO, Kolade AF, Nwanze JC. In-Vitro assessments of the effects of garlic (Allium sativum) extract on clinical isolates of Pseudomonas aeruginosa and Staphylococcus aureus. Adv Appl Sci Res. 2011;2(4): Ram Kumar Pundir, Pranay Jain, Chetan Sharma. Antimicrobial activity of ethanolic extracts of Syzygium aromaticum and Allium sativum. Ethnobotanical Leaflets. 2010;14: Bandna Chand. Antibacterial effect of garlic (Allium sativum). J of Microbiol, Biotech and Food Sci. 2013:2(4): Shoeb H. Antimicrobial susceptibility testing (Kirby Bauer) animation. Microbe Library, Available at: =2754&lang. Accessed 25 August Clinical and laboratory standards institute. Performans standards for antimicrobial susceptibility testing. Seventeenth Informational Supplement. 2007;CLSI 26:M100-S Srinivasan Durairaj, Sangeetha Srinivasan. In vitro antibacterial activity and stability of garlic extract at different ph and temperature. Electro J of Biol. 2009;5(1):5-10. DOI: / ijrms Cite this article as: Anitha M, Hemapriya J. Efficacy of antibacterial activity of garlic cloves from Tamil Nadu and Jowai region. Int J Res Med Sci 2014;2: International Journal of Research in Medical Sciences April-June 2014 Vol 2 Issue 2 Page 438